Proteases are widely used in a variety of contexts requiring the hydrolysis of proteins. It is possible to catagorize proteolytic hydrolysis processes into two general categories. One includes those procedures where the continuing presence of proteolytic activity after the accomplishment of the intended protein hydrolysis is of no concern. That category is conveniently typified by laundering operations with detergent formulations which contain proteases and by hide de-hairing processes employing protease formulations. In either case, the continuing presence of proteolytic activity is not material to the process.
A second category, and the context of concern in the present invention are those procedures where it is important to terminate proteolytic activity at some stage of the procedure, when a desired degree of protein hydrolysis is attained and when continuing activity would be undesirable or unacceptable.
The second category is typified by three convenient and commercially significant examples:
One is the protein hydrolysis of vegatable proteins, e.g. soy proteins, in order to produce a protein hydrolyzate with improved functional properties. Examples of such processes and the improvements in properties are set forth in U.S. Pat. Nos. 4,100,024 and 4,100,151, which are representative.
A second process is the protein hydrolysis carried out on animal proteins, such as the haemolyzed red blood corpuscle fraction of blood to obtain certain desired characteristics and properties in the hydrolyzate. A representative example of such a procedure is set out in Belgian Pat. No. 873,932.
A third process within the second category is the procedure employed in beer making known as chill-proofing which is in substance an enzymatic protein hydrolysis. Chill-proofing of beer is exemplified in U.S. Pat. No. 3,366,483.
In each of these processes, and in others of a like character, if proteolytic activity is still present after the conclusion of the intended degree of protein hydrolysis, the hydrolytic action will continue to a detrimental or undesired extent and begin to produce detrimental or undesired results. It is thus important to be able to deactivate the enzyme at the desired stage of the hydrolysis.
It is the ability to reliably and controllably deactivate proteolytic enzymes at the desired point in procedures such as those set forth above which is the object of the present invention.
The most usual technique for deactivating proteases is a heat treatment. Such heat treatments are simple and reliable, but are often accompanied by an unintended and detrimental degradation or denaturation of some proteins or protein hydrolyzates, which may unacceptably alter the properties or characteristics of the product.
Chemical deactivations have been developed, but are often limited in usefulness by one or more undesirable side effects or consequences thereof. Such techniques may, like heat treatments, degrade or denature the reaction products. They may also introduce undesirable or unacceptable by-products, such as toxic substances, adverse or off-taste components, or the like. In some cases chemical deactivation techniques may be prohibitively expensive.
It is accordingly an object of the present invention to provide Bacillus serine proteases which have been modified in such fashion that the thermal stability thereof is reduced to a level at which the proteolytic activity may be deactivated by a heat treatment under conditions sufficiently gentle that it will not degrade or denature the protein or protein hydrolyzate of the reaction product.